The health of the oral cavity is maintained by salivary secretions. The principal function of salivary glands is to produce these complex fluids. We utilize in vitro dispersed cells, and cultured epithelial cells of salivary glands, to understand mechanisms controlling saliva formation. We have focused these studies on autonomic neurotransmitter regulation of secretory events and associated signalling mechanisms. During this reporting period the primary focus of study has been directed at examining the role of GTP-binding regulatory (G) proteins in secretory stimuli in the rat parotid gland. In particular, we have shown that the Ca2+ mobilization response elicited by the muscarinic receptor agonist carbachol is unaffected by treatment with pertussis toxin (i.e. involves a pertussis toxin-insensitive G protein). We observed comparable carbachol sensitivity, and the ability of the agonist-sensitive intracellular Ca2+ pool to be refilled by extracellular Ca2+, in control and toxin-treated cells. Similarly, Ca2+ responses to AlF4 were unaffected in cells from pertussis toxin-treated rats. We also have characterized the high affinity GTPase activity in Gs-enriched parotid membranes. The beta-adrenergic receptor agonist isoproterenol can increase this GTPase activity (Vmax, about 5O%; no effect on Km) and stimulate cAMP formation in these membranes. Adenylyl cyclase activity is increased markedly in the presence of isoproterenol and GTPYS. Cholera toxin treatment of membranes decreases isoproterenol-induced high affinity GTPase activity and increases cAMP formation induced by GTP (+/- isoproterenol). These data suggest the agonist-induced increase in high affinity GTPase activity is involved in the deactivation of beta-adrenergic stimulated responses of rat parotid acinar cells.